New research

The Eureka Chain

Subtle changes in the membrane of a bacterial cell could help it evade the immune system; a new paper from TBSI researchers in Nature Communications explains its molecular structure and function

Jul 13, 2021Biochemistry, New Research

Postgrad Anna Hastings and the 800 MHz NMR spectrometer

Vincent Olieric (Paul Scherrer Institute), Samir Olatunji (TCD), and Chia-Ying Huang (Paul Scherrer Institute) at the Swiss Light Source signal victory in solving the first X-ray crystal structure of Lit.

By Trevor Butterworth

Tunderstand the biochemical structures and processes happening within cells at the scale of a few billionths of a meter requires a certain kind of fortitude. “It’s a very, very long process”, says Professor Martin Caffrey, Fellow Emeritus (Trinity and TBSI), “and when you start, you do not know whether there’s going to be an end until you get to the end”.

Three years ago, Caffrey’s lab at TBSI decided to analyze the molecular structure and processes of a recently discovered enzyme, lipoprotein intramolecular transacylase (Lit). This enzyme is involved in creating the lipoproteins that form part of the outer layer, the membrane, of a bacterial cell. As our immune systems target and lock onto these lipoproteins, understanding how the process by which they are created could help researchers develop new ways of treating bacterial infection.

There is a twist here. If you develop a therapy that kills the bacteria, you increase the evolutionary pressure on the bacteria to develop resistance—and anti-bacterial resistance is now a critical problem in healthcare. Finding ways to reduce the virulence of a bacterial infection without killing the bacteria takes the evolutionary pressure off. It’s a smarter, more sustainable approach to intervention.

Besides the importance and the intellectual excitement of doing fundamental research to describe new cellular structures and processes, this was an added incentive to take on the research and the risk.

Caffrey has devoted his life to understanding lipids and proteins in biological membranes. After completing a degree in Agriculture in UCD, he went in 1973 to Cornell University in Ithaca, New York, to study food science, which deepened his interest in chemistry, and then a PhD in biochemistry. After 20 years at Ohio State University, he returned to Ireland in 2002, first to the University of Limerick, and then to Trinity.

His research group has developed techniques to study the interactions between lipids and proteins in a cell membrane at an atomic level. To understand Lit, they would have to reduce it to a pure protein, take it out of the cell membrane, crystallize it, and hope the crystals diffracted at a high enough resolution to be interpretable.

“This is all very challenging”, says Caffrey. “You can fail at any step”. The first bacterium they tried produced very pure protein but didn’t crystalize. They switched to Bacillus cereus, a bacterium typically found in soil and food that can cause food poisoning. “We got these beautiful crystals, froze them with liquid nitrogen, and sent them off to Switzerland to be X-rayed”. This was the next point for potential failure.

“They have to diffract—the crystal needs to be ordered enough so that it will scatter the X-rays in a way such that the diffraction pattern can be back interpreted to give us the three-dimensional atomic blueprint of the protein—how the molecule is put together”, says Caffrey. In the case of a protein, it has thousands of atoms—so it is an extremely complicated process to come back from diffraction space into real space where we actually see atoms”.

But successful diffraction (see the moment the research team knew his happened in the photograph above) was only the start. They had to interpret what the blueprint meant—and then confirm that what was captured in the crystal reflected what was happening in the live membrane. To do this, they worked with a group who do molecular dynamic simulations at Queen’s University, Belfast.

It turns out that Lit shifts one of the two chains anchoring a protein in the membrane from one part to another of the terminal amino acid; in doing this, the bacteria’s ability to be sensed by the immune system decreases by a factor of 100. “Instead of the immune system launching and locking onto the bacteria”, says Caffrey, the bacteria are able to slink away, evading detection”.

These results have just been published in Nature Communications. They build on the work Caffrey’s research group has done on a related enzyme, lipoprotein signal peptidase II (LspA), which is now the subject of intense research as an antibiotic target.

“There were probably several eureka moments with Lit”, says Caffrey. “The most important result is that we now have an understanding of a new protein, a new activity and a reaction mechanism. The fundamentals are there”.

But they also have an intriguing hypothesis: Bacillus cereus and other pathogenic bacteria can evade the immune system by modifying its lipoproteins to be less immunogenic. To know if this hypothesis is valid will require engaging chemists, microbiologists and immunologists in follow-up testing.

“The beauty of TBSI”, says Caffrey, “is that we are surrounded by people like this”.

This research was funded by Science Foundation Ireland, the Irish Research Council, the German Research Foundation, the European Union’s Horizon 2020 research and innovation programme under the Marie-Skłodowska-Curie grants agreement, and the Northern Ireland Department of Agriculture, Environment and Rural Affairs.

Trevor Butterworth is an adjunct assistant professor of science communication in TBSI. He founded Sense About Science USA, is co-founder and VP of, and Chair of the Cardea steering group at Linux Foundation Public Health.

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